About Our Research
l Study on the mechanism of how RNA binding protein regulates gene alternative splicing in skin tumors and skin damage repair.
l The influence of anesthetic drugs on tumor occurrence and development.
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FAO：# as first author, * as corresponding author
Publications
Feiyu Long #, Liren Hu # , Yingxu Chen #, Xiaoxia Duan, Keliang Xie, Jianguo Feng*, Maohua Wang*. RBM3 is associated with acute lung injury in septic mice and patients via the NF-κB/NLRP3 pathway. Inflammation Research. 2023 Feb 13.
Sepsis refers to host response disorders caused by infection, leading to life-threatening organ dysfunction. RNA-binding motif protein 3 (RBM3) is an important cold-shock protein that is upregulated in response to mild hypothermia or hypoxia. In this study, we aimed to investigate whether RBM3 is involved in sepsis-associated acute lung injury (ALI).Intraperitoneal injection of LPS (10 mg/kg) was performed in wild type (WT) and RBM3 knockout (KO, RBM3−/−) mice to establish an in vivo sepsis model. An NLRP3 inflammasome inhibitor, MCC950 (50 mg/kg), was injected intraperitoneally 30 min before LPS treatment. Serum, lung tissues, and BALF were collected 24 h later for further analysis.
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Jianguo Feng*, Jianlong Zhou, Yunxiao Lin, Wenhua Huang*. hnRNP A1 in RNA metabolism regulation and as a potential therapeutic target. Frontiers in pharmacology. 2022 Oct 21;13:986409.
Abnormal RNA metabolism, regulated by various RNA binding proteins, can have functional consequences for multiple diseases. Heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) is an important RNA binding protein, that regulates various RNA metabolic processes, including transcription, alternative splicing of pre-mRNA, translation, miRNA processing and mRNA stability. As a potent splicing factor, hnRNP A1 can regulate multiple splicing events, including itself, collaborating with other cooperative or antagonistical splicing factors by binding to splicing sites and regulatory elements in exons or introns. hnRNPA1 can modulate gene transcription by directly interacting with promoters or indirectly impacting Pol II activities.
Shengfeng Deng#, Peng Yi#, Qian Yi*, Jianguo Feng*. Dysfunctional Gene Splicing in Glucose Metabolism may Contribute to Alzheimer’s Disease. Chinese Medical Journal. 2022 Jul 14.
The glucose metabolism is crucial for sustained brain activity as it provides energy and is a carbon source for multiple biomacromolecules; glucose metabolism decreases dramatically in Alzheimer’s disease (AD) and may be a fundamental cause for its development. Recent studies reveal that the alternative splicing events of certain genes effectively regulate several processes in glucose metabolism including Insulin receptor, insulin-degrading enzyme, pyruvate kinase M, receptor for Advanced Glycation Endproducts, and others, thereby, influencing glucose uptake, glycolysis, and advanced glycation end-products-mediated signaling pathways. Indeed, the discovery of aberrant alternative splicing that changes the proteomic diversity and protein activity in glucose metabolism has been pivotal in our understanding of AD development. In this review, we summarize the alternative splicing events of the glucose metabolism-related genes in AD pathology and highlight the crucial regulatory roles of splicing factors in the alternative splicing process. We also discuss the emerging therapeutic approaches for targeting splicing factors for AD treatment.
Xing Liu, Qinxue Hu, Qianxiu Chen, Jing Jia, Yong-Hong Liao*, Jianguo Feng*. Effect of dexmedetomidine for prevention of acute kidney injury after cardiac surgery: an updated systematic review and meta-analysis. Renal failure, 2022,44(1), 1150–1159.
Background: Acute kidney injury (AKI) is a serious complication related to cardiac surgery.Several studies have been conducted to investigate the effect of dexmedetomidine administration on AKI prevention.
Objective: To assess if dexmedetomidine is associated with a protective effect of renal function after cardiac surgery. And the aim of conducting this meta-analysis is to summarize the literature and determine the clinical utility of dexmedetomidine administration in patients undergoing cardiac surgery.
Methods: PubMed, Cochrane Library, and EMBASE databases were comprehensively searched for all randomized controlled trials (RCTs) published before 1 December, 2021 that investigated the effect of dexmedetomidine on AKI prevention.
Results: Our analysis included 16 studies involving 2148 patients. Compared with the control group, dexmedetomidine administration significantly reduced AKI incidence (OR, 0.47; 95% CI,0.36–0.61; p < 0.00001; I2 ¼ 26%) and the length of stay in the intensive care unit (ICU) but did not alter mortality rate, length of stay in the hospital, and mechanical ventilation time.Furthermore, the incidence of delirium among patients treated with dexmedetomidine was significantly decreased.
Conclusion: Dexmedetomidine administration has a positive effect on preventing AKI and post-operative delirium after cardiac surgery and significantly reduces the length of stay in the ICU.
Qian Yi#, Jianguo Feng#, Yi Liao*, Weichao Sun*. Circular RNAs in chemotherapy resistance of lung cancer and their potential therapeutic application. IUBMB Life.2022 May 20.
Lung cancer is one of the high malignancy-related incidence and mortality worldwide, accounting for about 13% of total cancer diagnoses. Currently, the use of anti-cancer agents is still the main therapeutic method for lung cancer. However, cancer cells will gradually show resistance to these drugs with the progress of treatment. And the molecular mechanisms underlying chemotherapy agents resistance remain unclear. circRNAs are newly identified noncoding RNAs molecules with covalently closed circular structures. Previous studies have shown that circRNAs are associated with tumorigenesis and progression of various cancers, including lung cancer. Recently, growing reports have suggested that circRNAs could contribute to drug resistance of lung cancer cell through different mechanisms. Therefore, in this review, we summarized the functions and underlying mechanisms of circRNAs in regulating chemoresistance of lung cancer and discussed their potential applications for diagnosis, prognosis, and treatment of lung cancer.
Qiuxia Wang, Jianguo Feng*, Liling Tang*.Non-Coding RNA Related to MAPK Signaling Pathway in Liver Cancer. International journal of molecular sciences, 2022,23(19), 11908.
The advancement in high-throughput sequencing analysis and the evaluation of chromatin state maps have revealed that eukaryotic cells produce many non-coding transcripts/RNAs. Further, a strong association was observed between some non-coding RNAs and cancer development. The mitogen-activated protein kinases (MAPK) belong to the serine–threonine kinase family and are the primary signaling pathways involved in cell proliferation from the cell surface to the nucleus. They play an important role in various human diseases. A few non-coding RNAs associated with the MAPK signaling pathway play a significant role in the development of several malignancies, including liver cancer. In this review, we summarize the molecular mechanisms and interactions of microRNA, lncRNA, and other non-coding RNAs in the development of liver cancer that are associated with the MAPK signaling pathway . Further, we briefly discuss the therapeutic strategies for liver cancer related to ncRNA and the MAPK signaling pathway
Xiang Huang#, Yirong Chen#, Junxiu Yi, Peng Yi, Jing Jia, Yonghong Liao, Jianguo Feng*, Xian Jiang*. Tetracaine hydrochloride induces cell cycle arrest in melanoma by downregulating hnRNPA1. Toxicology and Applied Pharmacology. 2022 Jan 1;434:115810.
Recent evidence suggests potential benefits of applying local anesthetics in cancer patients. Specifically, tetracaine has a potent antitumor effect in diverse cancers, including neuroblastoma, breast cancer, and melanoma; however, the underlying molecular mechanisms remain unclear. Here, we reported that tetracaine hydrochloride inhibited the growth of melanoma cells and arrested melanoma cells in the G0/G1 phase. Tetracaine hydrochloride treatment resulted in translocation of hnRNPA1 from the nucleoplasm to the nuclear envelope and reduced the protein stability of hnRNPA1 possibly by disrupting the dynamic balance of ubiquitination and neddylation. Elevated hnRNPA1 upregulated cyclin D1 to promote cell cycle in melanoma. The hnRNPA1 overexpression attenuated the effect of tetracaine hydrochloride on melanoma cell growth suppression and cell cycle arrest. Furthermore, melanoma homograft experiments demonstrated that tetracaine hydrochloride suppressed melanoma growth, while hnRNPA1 overexpression alleviated tetracaine’s antitumor effect on melanoma. Taken together, our findings suggest that tetracaine hydrochloride exerts a potent antitumor effect on melanoma both in vitro and in vivo, and the effect involves cell cycle arrest induction via downregulation of hnRNPA1.
Yi Liao#, Jianguo Feng#, Weichao Sun, Chao Wu, Jingyao Li, Tao Jing, Yuteng Liang, Yonghui Qian, Wenlan Liu*, Haidong Wang*. CIRP promotes the progression of non-small cell lung cancer through activation of Wnt/β-catenin signaling via CTNNB1. Journal of Experimental & Clinical Cancer Research. 2021 Aug 31;40(1):275.
Background: Cold-inducible RNA binding protein (CIRP) is a newly discovered proto-oncogene. In this study, we investigated the role of CIRP in the progression of non-small cell lung cancer (NSCLC) using patient tissue samples, cultured cell lines and animal lung cancer models.
Methods: Tissue arrays, IHC and HE staining, immunoblotting, and qRT-PCR were used to detect the indicated gene expression; plasmid and siRNA transfections as well as viral infection were used to manipulate gene expression; cell proliferation assay, cell cycle analysis, cell migration and invasion analysis, soft agar colony formation assay, tail intravenous injection and subcutaneous inoculation of animal models were performed to study the role of CIRP in NSCLC cells; Gene expression microarray was used to select the underlying pathways; and RNA immunoprecipitation assay, biotin pull-down assay, immunopurification assay, mRNA decay analyses and luciferase reporter assay were performed to elucidate the mechanisms. The log-rank (Mantel-Cox) test, independent sample T-test, nonparametric Mann-Whitney test, Spearman rank test and two-tailed independent sample T-test were used accordingly in our study.
Results: Our data showed that CIRP was highly expressed in NSCLC tissue, and its level was negatively correlated with the prognosis of NSCLC patients. By manipulating CIRP expression in A549, H460, H1299, and H1650 cell lines, we demonstrated that CIRP overexpression promoted the transition of G1/G0 phase to S phase and the formation of an enhanced malignant phenotype of NSCLC, reflected by increased proliferation, enhanced invasion/metastasis and greater tumorigenic capabilities both in vitro and in vivo. Transcriptome sequencing further demonstrated that CIRP acted on the cell cycle, DNA replication and Wnt signaling pathway to exert its pro-oncogenic action. Mechanistically, CIRP directly bound to the 3′- and 5′-UTRs of CTNNB1 mRNA, leading to enhanced stability and translation of CTNNB1 mRNA and promoting IRES-mediated protein synthesis, respectively. Eventually, the increased CTNNB1 protein levels mediated excessive activation of the Wnt/β-catenin signaling pathway and its downstream targets C-myc, COX-2, CCND1, MMP7, VEGFA and CD44.
Mengting Ou, Xichao Xu, Ying Chen, Li Li, Lu Zhang, Yi Liao, Weichao Sun, Christine Quach, Jianguo Feng*, Liling Tang*. MDM2 induces EMT via the B‑Raf signaling pathway through 14‑3‑3. Oncol Rep. 2021 Jul;46(1):120.
MDM2 proto‑oncogene, E3 ubiquitin protein ligase (MDM2) is a well‑known oncogene and has been reported to be closely associated with epithelial‑to‑mesenchymal transition (EMT). The present study first demonstrated that the expression levels of MDM2 were markedly increased in TGF‑β‑induced EMT using quantitative PCR and western blotting. In addition, MDM2 was demonstrated to be associated with pathological grade in clinical glioma samples by immunohistochemical staining. Furthermore, overexpression of MDM2 promoted EMT in glioma, lung cancer and breast cancer cell lines using a scratch wound migration assay.
Jianguo Feng#, Xichao Xu#, Xin Fan, Qian Yi, Liling Tang*. BAF57/SMARCE1 interacting with Splicing factor SRSF1 regulates mechanical stress-induced alternative splicing of cyclin D1. Genes, 2021, 12(2): 306.
Background: Cyclin D1 regulates cyclin-dependent protein kinase activity of the cell cycle, and cyclin D1 alternative splicing generates a cyclin D1b isoform, acting as a mediator of aberrant cellular proliferation. As alternative splicing processes are sensitive to mechanical stimuli, whether the alternative splicing of cyclin D1 is regulated by mechanical stress and what kinds of factors may act as the regulator of mechano-induced alternative splicing remain unknown.
Methods: The alternative splicing of Cyclin D1 was examined using reverse transcription polymerase chain reaction (RT-PCR) in osteoblast cell lines and keratinocyte cells loaded by a cyclic stretch. The expression of splicing factors and switching defective/sucrose non-fermenting (SWI/SNF) complex subunits were detected in stretched cells using real-time quantitative PCR (RT-qPCR). The protein interaction was tested by co-immunoprecipitation assay (Co-IP).
Results: Cyclin D1 expression decreased with its splice variant upregulated in stretched cells. Serine/arginine-rich splicing factor 1(SRSF1) and SWI/SNF complex subunit Brahma-related gene-1-associated factor 57 (BAF57), also named SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily Emember 1 (SMARCE1), could respond to mechanical stimuli. Overexpression and knockdown experiments indicated the BAF57/SMARCE1 is probably a critical factor regulating the alternative splicing of cyclin D1. Co-IP showed an interaction between BAF57/SMARCE1 and SRSF1, implying a possible underlying mechanism of the regulator role of BAF57/SMARCE1 in the splicing process of cyclin D1.
Conclusions: The splicing factor SRSF1 and BAF57/SMARCE1 are possibly responsible for the mechanical stress-induced alternative splicing of cyclin D1.
Xin He#, Ainong Zhu#, Jianguo Feng*, Xiaobin Wang*. Role of neddylation in neurological development and diseases. Biotechnology and Applied Biochemistry. 2022 Feb;69(1):330-341.
Neddylation, a posttranslational protein modification, refers to the specific conjugation of NEDD8 to substrates, which is of great significance to various biological processes. Besides members of the cullin protein family, other key proteins can act as a substrate for neddylation modification, which remarkably influences neurodevelopment and neurodegenerative diseases. Normal levels of protein neddylation contribute to nerve growth, synapse strength, neurotransmission, and synaptic plasticity, whereas overactivation of protein neddylation pathways lead to apoptosis, autophagy of neurons, and tumorigenesis. Furthermore, impaired neddylation causes neurodegenerative diseases. These facts suggest that neddylation may be a target for treatment of these diseases. This review focuses on the current understanding of neddylation function in neurodevelopment as well as neurodegenerative diseases. Meanwhile, the recent view that different level of neddylation pathway may contribute to the opposing disease progression, such as neoplasms and Alzheimer’ s disease, is discussed. The review also discusses neddylation inhibitors, which are currently being tested in clinical trials. However, potential drawbacks of these drugs are noted, which may benefit the development of new pharmaceutical strategies in the treatment of nervous system diseases.
Rongxue Wan, Xichao Xu, Lunkun Ma, Ying Chen, Liling Tang*, Jianguo Feng*. Novel Alternatively Spliced Variants of Smad4 Expressed in TGF-β-Induced EMT Regulating Proliferation and Migration of A549 Cells. Onco Targets Ther. 2020;11;13:2203-2213.
Introduction: Non-small cell lung cancer (NSCLC) is a worldwide malignance threatening human life. TGF-β/Smad signaling is known to regulate cell proliferation, differentiation, migration and growth. As the only co-Smad playing crucial roles in TGF-β signaling, Smad4 is reported to be frequently mutated or to occur as alternatively spliced in tumor cells. Smad4 was reported to be involved in the TGF-β-induced EMT process. However, whether the alternative splicing occurs in the TGF-β-induced EMT process in NSCLC was not clear.
Methods: In our current study, we explored the alternative splicing of Smad4 during the process of TGF-β-induced EMT in A549 cells. 10 ng/mL TGF-β was used to induce EMT. Then, nest-PCR and agarose electrophoresis were performed to detect the expression of Smad4 variants and sequencing to get the variant DNA sequences. For recombinant expression of variants of Smad4 in A549 cells, we used lentiviral variants to infect cells. In order to explore the effects of variants on the proliferation and migration of A549 cells, the MTT assay, colony formation assay and wound-healing assay were done. The effects of variants on E-cad and VIM protein expression were explored through Western blot.
Results: There were several novel gene fragments expressed in TGF-β-induced A549 cells, and the sequencing results showed that they were indeed the Smad4 variants that were not reported. For recombinant expression of Smad4 variants in A549 cells, we found that they have significant effects on the proliferation and migration of cells, and also regulated the E-cad and VIM protein expression.
Conclusion: Our results indicated that novel Smad4 variants were expressed in TGF-β- induced EMT process. The functional study showed that these novel variants regulate cell proliferation and migration and affect E-cad and VIM protein expression, showing the potential as targets for cancer therapy.
Meilian Zhao, Jianguo Feng*, Liling Tang*. Competing Endogenous RNAs in Lung Cancer. Cancer Biology & Medicine 2021 Feb 15;18(1):1-20.
Competing endogenous RNAs (ceRNAs) containing microRNA response elements can competitively interact with microRNA via miRNA response elements, which can combine non-coding RNAs with protein-coding RNAs through complex ceRNA networks. CeRNAs include non-coding RNAs (long non-coding RNAs, circular RNAs, and transcribed pseudogenes) and protein-coding RNAs (mRNAs). Molecular interactions in ceRNA networks can coordinate many biological processes; however, they may also lead to ceRNA network imbalance and thus contribute to cancer occurrence when disturbed. Recent studies indicate that many dysregulated RNAs derived from lung cancer may function as ceRNAs to regulate multitudinous biological functions for lung cancer, including tumor cell proliferation, apoptosis, growth, invasion, migration, and metastasis. This study therefore reviewed the research progress in the field of non-coding and protein-coding RNAs as ceRNAs in lung cancer, and highlighted validated ceRNAs involved in biological lung cancer functions. Furthermore, the roles of ceRNAs as novel prognostic and diagnostic biomarkers were also discussed. Interpreting the involvement of ceRNAs networks in lung cancer will provide new insight into cancer pathogenesis and treatment strategies.
Huifang Sun, Jianguo Feng*, Liling Tang*. Function of TREM1 and TREM2 in Liver-Related Diseases. Cells. 2020,7;9(12):E2626. doi: 10.3390/cells9122626.
TREM1 and TREM2 are members of the triggering receptors expressed on myeloid cells (TREM) family . Both TREM1 and TREM2 are immunoglobulin superfamily receptors. Their main function is to identify foreign antigens and toxic substances, thereby adjusting the inflammatory response. In the liver, TREM1 and TREM2 are expressed on non-parenchymal cells, such as liver sinusoidal endothelial cells, Kupffer cells, and hepatic stellate cells, and cells which infiltrate the liver in response to injury including monocyte-derived macrophages and neutrophils. The function of TREM1 and TREM2 in inflammatory response depends on Toll-like receptor 4. TREM1 mainly augments inflammation during acute inflammation, while TREM2 mainly inhibits chronic inflammation to protect the liver from pathological changes. Chronic inflammation often induces metabolic abnormalities, fibrosis, and tumorigenesis. The above physiological changes lead to liver-related diseases, such as liver injury, nonalcoholic steatohepatitis, hepatic fibrosis, and hepatocellular carcinoma. Here, we review the function of TREM1 and TREM2 in different liver diseases based on inflammation, providing a more comprehensive perspective for the treatment of liver-related diseases.
Jianguo Feng#, Maozhou Wang#, Mao Li, Jimei Yang, Jing Jia, Li Liu, Jun Zhou, Chunxiang Zhang, Xiaobin Wang*. Serum miR-221-3p as a new potential biomarker for depressed mood in perioperative patients. Brain Res. 2019;1720:146296.
MicroRNAs (miRNAs) modulate various genes associated with brain disorders and circulating miRNAs may therefore serve as biomarkers for these neurological diseases. We previously found that the miRNA miR-221-3p was highly expressed in cerebrospinal fluid and the serum of major depressive disorder (MDD) patients. Here, we examined whether miR-221-3p could be used as a biomarker for depressed mood in perioperative patients. We first examined the relative expression of serum miR-221-3p by real-time quantitative PCR in perioperative patients with different degrees of depressive mood assessed by the Patient Health Questionnaire-9 (PHQ-9) diagnostic form. We found that miR-221-3p expression in the mild depressive mood group (PHQ-9 scores 5–9) was 2.21 fold that of the normal group (PHQ-9 scores 0–4) and the moderate＆severe depressive mood group (PHQ-9 scores ≥ 10) showed miR-221-3p expression levels 3.66 fold that of the normal group. Then the absolute quantification of serum miR-221-3p was obtained using an miRNA standard curve. We found that the amount of serum miR-221-3p was positively correlated with depressed mood; when serum miR-221-3p > 1.7 × 107 copies/μg RNA, all indicated PHQ-9 scores were higher than 6. Subsequently, we found that miR-221-3p could indirectly increase the expression of IFN-α (Interferon alpha) in astrocytes by targeting IRF2(Interferon Regulatory Factor 2) and that miR-221-3p participated in the anti-neuroinflammatory signaling cascades induced by ketamine and paroxetine via the IRF2/IFN-α pathway. Our results indicate that elevated serum miR-221-3p can be used as a biomarker for depressed mood in perioperative patients and that IFN-α-induced NF-κB activation in astrocytes mediated by miR-221-3p targeting of IRF2 may be one of the potential mechanisms.
Irfan Ullah, Yi Liao, Rongxue Wan, Liling Tang*, Jianguo Feng*. Alternative Splicing of SMAD4 and Its Function in HaCaT Cells in Response to UVB Irradiation. J Cancer 2018; 9(17):3177-3186.
Alternative splicing is one of the most common mechanisms of human gene regulation and plays a crucial role in increasing the diversity of functional proteins. Many diseases are linked to alternative splicing, especially cancer. SMAD4 is a member of the SMAD family and plays a critical role in mediating of TGF-β signal transduction and gene regulatory events. Smad4 is a tumour suppressor and acts as a shuttling protein between nucleus and cytoplasm. The splicing variants of Smad4 have been found in many cancers. The present study performed nested PCR to detect alternative splicing of Smad4 in HaCaT cells lines in response to UVB irradiation. The UVB induced a novel Smad4B isoform that led to decrease the Smad4 expression. The hnRNPA1 splicing factor is responsible for Smad4 alternative splicing in response to UVB. The UVB increased the expression of SF2 and hnRNPA1 Splicing factors. The hnRNPA1 overexpression induced Smad4B by regulating Smad4 alternative splicing. The Smad4B isoform supported the function of Smad4 full length in UVB resistance with certain limitation. The western blot analyses showed that the overexpressed Smad4 full length significantly increased N-cadherin expression while Smad4B overexpression decreased the expression the N-cadherin (P<0.05). Furthermore, overexpression of the isoform in HaCaT cells decreased cell invasion as compared to Smad4 full-length overexpression. These results will be helpful to understand the importance of Smad4 alternative splicing in skin tumorigenesis.
Irfan Ullah, Weichao Sun, Liling Tang*, Jianguo Feng*. Roles of Smads Family and Alternative Splicing Variants of Smad4 in Different Cancers. J Cancer, 2018; 9(21): 4018-4028.
Transforming Growth Factor β (TGF-β) is one of the most common secretory proteins which are recognized by membrane receptors joined to transcription regulatory factor. TGF-β signals are transduced by the Smads family that regulate differentiation, proliferation, early growth, apoptosis, homeostasis, and tumor development. Functional study of TGF-β signaling pathway and Smads role is vital for certain diseases such as cancer. Alternative splicing produces a diverse range of protein isoforms with unique function and the ability to react differently with various pharmaceutical products. This review organizes to describe the general study of Smads family, the process of alternative splicing, the general aspect of alternative splicing of Smad4 in cancer and the possible use of spliceoforms for the diagnosis and therapeutic purpose. The main aim and objective of this article are to highlight some particular mechanisms involving in alternatives splicing of cancer and also to demonstrate new evidence about alternative splicing in different steps given cancer initiation and progression.
Yun-Qiang Wan#, Jian-Guo Feng#, Mao Li#, Mao-Zhou Wang, Li Liu, Xueru Liu, Xiao-Xia Duan, Chun-Xiang Zhang, Xiao-Bin Wang*. Prefrontal cortex miR-29b-3p plays a key role in the antidepressant-like effect of ketamine in rats. Experimental & Molecular Medicine 2018 Oct 29;50(10):140.
Ketamine has a rapid, obvious, and persistent antidepressant effect, but its underlying molecular mechanisms remain unknown. Recently, microRNAs (miRNAs) have emerged as important modulators of ketamine’s antidepressant effect. We investigated the alteration in miR-29b-3p in the brain of rats subjected to ketamine administration and chronic unpredictable mild stress (CUMS), and a sucrose preference test and forced swimming test were used to evaluate the rats’ depressive-like state. We used recombination adeno-associated virus (rAAV) or lentivirus-expressing miR-29b-3p to observe the change in metabotropic glutamate receptor 4 (GRM4). Cell culture and electrophysiological recordings were used to evaluate the function of miR-29b-3p. Ketamine dramatically increased miR-29b-3p expression in the prefrontal cortex of the normal rats. The dual luciferase reporter test confirmed that GRM4 was the target of miR-29b-3p. The miR-29b-3p levels were downregulated, while the GRM4 levels were upregulated in the prefrontal cortex of the depressive-like rats. The ketamine treatment increased miR-29b-3p expression and decreased GRM4 expression in the prefrontal cortex of the depressive-like rats and primary neurons. By overexpressing and silencing miR-29b-3p, we further validated that miR-29b-3p could negatively regulate GRM4. The silencing of miR-29b-3p suppressed the Ca2+ influx in the prefrontal cortex neurons. The miR-29b-3p overexpression contributed to cell survival, cytodendrite growth, increases in extracellular glutamate concentration, and cell apoptosis inhibition. The overexpression of miR-29b-3p by rAAV resulted in a noticeable relief of the depressive behaviors of the CUMS rats and a lower expression of GRM4. The miR-29b-3p/GRM4 pathway acts as a critical mediator of ketamine’s antidepressant effect in depressive-like rats and could be considered a potential therapeutic target for treating major depression disorder.
Feng Jianguo#, Liao Yi#, Xu Xichao, Yi Qian, He Ling, Tang Liling*. hnRNP A1 promotes keratinocyte cell survival post UVB radiation through PI3K/Akt/mTOR pathway. Exp Cell Res. 2018, 15;362(2):394-399.
hnRNP A1 acts as a critical splicing factor in regulating many alternative splicing events in various physiological and pathophysiological progressions. hnRNP A1 is capable of regulating UVB-induced hdm2 gene alternative splicing according to our previous study. However, the biological function and underlying molecular mechanism of hnRNP A1 in cell survival and cell cycle in response to UVB irradiation are still unclear. In this study, silencing hnRNP A1 expression by siRNA transfection led to decreased cell survival after UVB treatment, while promoting hnRNP A1 by lentiviruse vector resulted in increased cell survival. hnRNP A1 remarkably enhanced PI3K/Akt/mTOR signaling pathway by increasing phosphorylation of Akt, mTOR and P70S6 protein. Inhibition of PI3K/Akt signaling by LY294002 suppressed the expression of hnRNP A1. While mTOR signaling inhibitors, rapamycin and AZD8055, did not influence hnRNP A1 expression in HaCaT cells, suggesting that hnRNP A1 may be an upstream mediator of mTOR signaling. Furthermore, hnRNP A1 could alleviate UVB-provoked cell cycle arrest at G0/G1 phase and promoted cell cycle progression at G2/M phase. Our results indicate that hnRNP A1 promotes cell survival and cell cycle progression following UVB radiation.
Liao Yi#, Feng Jianguo#, Zhang Yi, Tang Liling*, Wu Shiyong*.The mechanism of CIRP in inhibition of keratinocytes growth arrest and apoptosis following low dose UVB radiation. Mol Carcinog. 2017，56(6):1554-1569.
UV induces CIRP expression and subsequent Stat3 activation, but the biological function and mechanism of CIRP and Stat3 in mediating UVB-induced skin carcinogenesis have not been fully elucidated. In this study, we demonstrate that CIRP is elevated in all tested melanoma and non-melanoma skin cancer cell lines; and the expression of CIRP is upregulated in keratinocytes after being irradiated with relatively low dose (<5 mJ/cm2), but not high dose (50 mJ/cm2), UVB acutely and chronically. The increased expression of CIRP, either induced by UVB or through overexpression, leads to resistance of keratinocytes to UVB-induced growth arrest and death; and reduced expression of CIRP by RNA knockdown sensitizes keratinocyte cells to the low dose UVB radiation. We also demonstrated that CIRP expression is required for the low dose UVB-induced Tyr705-phosphorylation, but not total amount, of Stat3. The p-Stat3 level is correlated with the expression levels of cyclin D1 and VEGF, two known downstream cell growth regulators of Stat3, as well as Bag-1/S, an apoptosis regulator. Inhibition of Stat3 DNA-binding activity by S3I-201 leads to a reduction of the p-Stat3 and Bag-1/S along with growth and survival of keratinocytes post-UVB; and the effect of S3I-201 on the UVB-irradiated cells can be partially inhibited by overexpression of CIRP or Bag-1/S. Furthermore, the overexpression of Bag-1/S can totally inhibit UVB-induced PARP cleavage and caspase 3 activation. The results presented above led us to propose that CIRP-p(705)Stat3 cascade promotes cell proliferation and survival post-UVB via upregulating the expression of cyclin D1 and Bag-1/S, respectively. Published 2017. This article is a U.S. Government work and is in the public domain in the USA.
Jianguo Feng, Li Li, Lingying Tong, Liling Tang*, Shiyong Wu*. The Involvement Of Splicing Factor hnRNP A1 In UVB-Induced Alternative Splicing Of hdm2. Photochem Photobiol. 2016,92(2):318-324.
Human homolog double minute 2 (hdm2), an oncoprotein, which binds to tumor suppressor p53 to facilitate its degradation, has been known to contribute to tumorigenesis. Its splicing variants are reported to be highly expressed in many cancers and can be induced by ultraviolet B light (UVB). However, the mechanisms of how UVB radiation induces hdm2 alternative splicing still remain unclear. In this study, we investigated the roles of two common splicing factors, heterogeneous nuclear ribonucleoproteins (hnRNP) A1 and serine/arginine-rich splicing factor 1 (SRSF1), in regulating UVB-induced hdm2 splicing. Our study indicated that while the expression of both hnRNP A1 and SRSF1 are induced, only hnRNP A1 is involved in hdm2 alternative splicing upon UVB irradiation. Overexpression of hnRNP A1 resulted in decrease of full-length hdm2 (hdm2-FL) and increase of hdm2B, one of hdm2 alternate-splicing forms; while down-regulated hnRNP A1 expression led to the decrease of the hdm2-FL and hdm2B in HaCaT cells. Protein-mRNA binding assay confirmed that UVB irradiation could increase the binding of hnRNP A1 to hdm2 pre-mRNA. In conclusion, we elucidated that UVB induces alternative splicing of hdm2 via increasing the expression and the binding of hnRNP A1 to hdm2 full-length mRNA.
Jianguo Feng，Rongxue Wan, Qian Yi, Ling He, Li Yang, Liling Tang*. Examination of Alternate Codon Bias Solutions for Expression for expression and purification of recombinant mechano growth factor in Escherichia coli. Biotechnol Appl Biochem. 2015;62(5):690-698.
Mechano-growth factor (MGF), an alternative splicing variant of insulin-like growth factor-1 (IGF-1) gene, promotes cell proliferation and inhibits cell differentiation. It also plays an important role in tumor development. It is important to optimize the production process and achieve MGF protein because there is no commercial MGF protein available. In this study, the human MGF gene is cloned into pGEX-4T-1 and the recombinant human MGF (rhMGF) protein could be expressed in Rosetta (DE3) by isopropyl β-D-1-thiogalactopyranoside induction but not in BL21 (DE3). Mutation from rare codons to Escherichia coli preferred ones is performed. We obtain MGF(Mut54–56) and MGF(Mut-total) fragments through site-directed mutagenesis and overlapping PCR. Both pGEX-4T-1/MGF(Mut54–56)- and pGEX-4T-1/MGF(Mut-total)-transformed BL21 (DE3) can be induced to express rhMGF protein. T o optimize the production technology, expression and purification of rhMGF are analyzed and compared in Rosetta (DE3) and BL21 (DE3). Results indicate that rhMGF expression in BL21 (DE3) is significantly higher than that in Rosetta (DE3). The protein yield of pGEX-4T-1/MGF(Mut-total) in BL21 (DE3) is higher than that of pGEX-4T-1/MGF(Mut54–56). We test the biological activity of MGF protein purified by affinity chromatography in C2C12 cell line and find that rhMGF promotes cell proliferation significantly. In conclusion, we establish a method to produce rhMGF economically with high biological activity in BL21 (DE3).
Jianguo Feng#, Wei Pan#, Xiaoli Yang, Feiyu Long, Jun Zhou, Yi Liao*, Maohua Wang*. RBM3 Increases Cell Survival but Disrupts Tight Junction of Microvascular Endothelial Cells in Acute Lung Injury. Journal of Surgical Research. 2021 Jan 15;261:226-235.
Background: RNA-binding motif protein 3 (RBM3) is an important cold shock protein, which also responds to hypothermia or hypoxia. RBM3 is involved into multiple physiologic processes, such as promoting cell survival. However, its expression and function in acute lung injury (ALI) have not been reported.
Methods:A mouse ALI model was established by lipopolysaccharides (LPS) treatment. The RBM3 and cold inducible RNA-binding protein mRNA levels were examined by RT-qPCR, and MMP9 mRNA stability was determined by actinomycin D assay. RBM3 and MMP9 mRNA was tested by RNA immunoprecipitation (RIP assay). RBM3 overexpression or silent stable cell lines were established using recombinant lentivirus and subsequently used for cell survival and tight junction measurements.
Results:In this study, we found that RBM3, rather than cold inducible RNA-binding protein, was upregulated in lung tissue of ALI mice. RBM3 was increased in human pulmonary microvascular endothelial cells (HPMVECs) in response to LPS treatment, which is modulated by the NF-κB signaling pathway. Furthermore, RBM3 could reduce cell apoptosis induced by LPS, probably through suppressing p53 expression. Because increased permeability of HPMVECs leads to pulmonary edema in ALI, we subsequently examined the effect of RBM3 on cell tight junctions. Unexpectedly, RBM3 decreased the expression of tight junction protein zonula occludens-1 and increased cell permeability, and RBM3 overexpression increased MMP9 mRNA stability. Furthermore, RIP assay confirmed the interaction between RBM3 and MMP9 mRNA, possibly explaining the contribution of RBM3 to increase cell permeability.
Conclusions:RBM3 seems to act as a “double-edged sword” in ALI, that RBM3 alleviates cell apoptosis but increases HPMVEC permeability in ALI.
Jianguo Feng, Liling Tang*. SPARC in Tumor Pathophysiology and as a Potential Therapeutic Target. Curr Pharm Des. 2014;20:6182-6190.
Cell migration and metastasis greatly contribute to the progression of tumors. Secreted Protein and Rich in Cysteine （SPARC）, as a multi-faceted protein, is highly expressed in highly metastatic tumors while low or undetectable in less metastatic types with aberrant promoter methylation. In highly metastatic tumors, such as glioblastomas, melanoma, breast cancer and prostate cancer, SPARC promotes bone metastasis and epithelial-mesenchymal transition (EMT). In contrast, this protein acts as an anti-tumor factor in anti-angiogenesis, pro-apoptosis, cell proliferation inhibition and cell cycle arrest in less metastatic tumors, such as neuroblastoma, ovarian cancer, pancreatic cancer, colorectal cancer and gastric cancer. Here, we summarize and analyze the paradoxical role of SPARC in different tumors. We believe that further studies on truncated, alternative splicing variants and signal peptide of SPARC are required to elucidate the distinct effects. Most notably, SPARC variants probably play a crucial role in regulation of transforming growth factor beta (TGF-β) induced EMT. This review also provides strategies to target or use SPARC (full-length, truncated and splicing variants) for therapeutic purposes.